Suppressor of cytokine signaling (SOCS) proteins were discovered as negative regulators of the cytokine receptor signaling pathways. Our long-term goal is to define the role of the IGF system in development of human cancers and to characterize downsteam regulatory components of this important signaling system. The primary goal of this application is to define the role of SOCS-3 in IGF-IR signal transduction in colon cancer. We have identified human SOCS-3 as an intracellular binding partner of the activated insulin-like growth factor-I receptor (IGF-IR). SOCS-3 interacts with the IGF-IR cytoplasmic domain both in vitro and in vivo in mammalian cells. The IGF-IR plays an crucial role in promoting the growth of cancers by stimulating malignant cell proliferation and transformation, and by inhibiting apoptosis. In human colon cancer Caco-2 cells, the IGF-IR and IGF-II are expressed at high levels and SOCS-3 is significantly induced by IGF-I stimulation. SOCS-3 is phosphorylated on tyrosine residues as a result of its interaction with the activated IGF-IR and thus is a substrate of the IGF-IR. SOCS-3 is heavily phosphorylated at serine residues and is ubiquitinated in mammalian cells. SOCS-3 protein is localized mostly in the cytoplasm but IGF-I stimulation results in translocation to the nucleus. SOCS-3 mRNA is expressed in many human fetal and adult human tissues, and some human cancer cell lines. Very little is known about the mechanism of SOCS-3 action. We hypothesize that SOCS-3 protein plays a negative regulatory role in IGF-IR signal transduction in colon cancer and we have formulated four specific aims to test this hypothesis. We propose to: 1) Determine the domain(s) of SOCS-3 responsible for IGF-IR interaction; 2) Identify the binding site/motif for SOCS-3 protein within the cytoplasmic domain of the IGF-IR; 3) Determine which signal transduction pathway(s) downstream of the IGF-IR (e.g. MAP kinase, PI 3-kinase, or others) are responsible for induction of SOCS-3 in Caco-2 cells and determine if SOCS-3 inhibits IGF-I induced mitogenic signaling in these cells; 4) Determine if IGF-IR-SOCS-3 interactions interfere with IGF-IR signaling in Caco-2 cells by using siRNA to silence SOCS-3 gene expression. We will explore the possibility of negative regulation of the IGF-IR signaling by SOCS-3 as a new example of growth resistance in cancer.